SECONDA CONFERENZA DEI PROGETTI DEL CENTRO FERMI APRILE 2012 SALONE DELLE CONFERENZE DEL MINISTERO DELLINTERNO ROMA, PIAZZA DEL VIMINALE 1 In vitro-In vivo MUrine coSmiC siLEnce experiment (MUSCLE) D. Capece and E. Fratini (Centro Fermi) on behalf of the Cosmic Silence Collaboration

Aim of MUSCLE Project To understand the biological effects of environmental radiation To understand the biological effects of environmental radiation

Peculiar low dose radiation effects (<100mGy) Phenomenon in which unirradiated cells exhibit irradiated effects as a result of signals received from nearby irradiated cells. Bystander effect

Genomic instability Phenomenon characterized by an increase in the genetic spontaneous alteration observed in cells which have not themselves been irradiated, but are the progeny of an irradiated cell. Peculiar low dose radiation effects (<100mGy)

Conditioning DoseChallenge Dose Damage + Peculiar low dose radiation effects (<100mGy) Adaptive reponse Phenomenon that occurs when a low dose of mutagenic agent can protect from the biological damage of a subsequent high dose of the same or different mutagenica agents

Our Hypothesis Life has evolved on Earth for 3 bilion years in the presence of environmental ionizing radiation. All organism have therefore incorporated in their normal biology a daily stiimulus of ultra-low-dose environmental radiation that might act as conditioning agent.

LNGS – The Gran Sasso underground laboratory

Past Experiments S. Cerevisiae cells Chinese Hamster cells Human cells Satta et al., Satta et al., Antonelli et al., Carbone et al., Carbone & Pinto et al., 2010.

S. Cerevisiae D7 cells Satta et al., Mutat. Res

Chinese Hamster V79 Satta et al., Radiat. Environ. Biophys Antonelli et al., Nuovo Cimento B 2008.

TK6 Lymphoblastoid cells Carbone et al., Radiat. Environ. Biophys Carbone & Pinto et al., Nuovo Cimento B 2010.

…Summarizing… Compatible with our hypothesis of environmental radiation as a conditioning agent, reduced environmental radiation conditions seem to render cells, after several months of protracted culture: Less tolerant to radiation-induced DNA damage Less efficient in scavenging reactive oxygen species Compatible with our hypothesis of environmental radiation as a conditioning agent, reduced environmental radiation conditions seem to render cells, after several months of protracted culture: Less tolerant to radiation-induced DNA damage Less efficient in scavenging reactive oxygen species

pKZ1 Mouse Model An ideal model for studying responses to very low doses of DNA damaging agents Matsuoka et al., Science 1991

LacZ Expression Matsuoka et. al., Science 1991 pKZ1 Chromosomal Inversion Assay P P P P DNA Inversion ZcaL LacZ

Experimental design A11 Hybridoma cell line obtained from pKZ1 a splenocyte Immortalized cell lines from brain and skin of neonatal pKZ1 mice To use pKZ1 mouse chromosomal inversion assay to study biological effects of environmental background radiation Day et al., Radiat. Res. 1991

Experimental design Istituto Superiore di Sanità Rome LNGS - INFN Reference environmental radiation (~ 2mGy/year) Reduced environmental radiation (~ 25mGy/year) Environmental Radiation Compnents ISS (External/Reference Laboratory) LNGS (Underground Laboratory) Cosmic Rays30 nGy/hNegligible Cosmic and terrestrial - rays 300 nGy/h3.6 nGy/h 222 Rn1.7 nGy/h0.17 nGy/h Total dose rate~ 330 nGy/h~ 3.8 nGy/h

Experimental design RomeTeramo Inversion frequency Doubling Time Rate of apoptosis/autophagy Redox cellular state Biological End-points ISS – Reference Environmental Radiation LNGS – Reduced Environmental Radiation

Reference Environmental Radiation LAquila University Reduced Environmental Radiation LNGS (LAquila)/LSC-Canfranc underground laboratory (Saragozza)? pKZ1 Perspectives What happens in vivo ?

INFN L. Satta Istituto Superiore di Sanità G. Simone, M.A. Tabocchini LAquila University F. Zazzeroni, E. Alesse ENEA Casaccia R. Amendola INFN-LNGS M. Balata ENEA R. Amendola Flinders University, Adelaide P. Sykes Cosmic Silence Collaboration CENTRO FERMI D. Capece, E. Fratini, M.C. Carbone, M. Pinto